Sienes Bailo et al. BMC Medical Genetics(2020) 21:233
https://doi.org/10.1186/s12881-020-01174-z

RESEARCH ARTICLE

Open Access

Incidence of Huntington disease in anortheastern Spanish region: a 13-year
retrospective study at tertiary care centre
Paula Sienes Bailo, Raquel Lahoz* , Juan Pelegrín Sánchez Marín and Silvia Izquierdo Álvarez

Abstract
Background: Despite the progress in the knowledge of Huntington disease (HD) in recent years, the epidemiology
continues uncertain, so the study of incidence becomes relevant. This is important since various factors (type of
population, diagnostic criteria, disease-modifying factors, etc.) make these data highly variable. Therefore, thegenetic diagnosis of these patients is important, since it unequivocally allows the detection of new cases.
Methods: Descriptive retrospective study with 179 individuals. Incidence of HD was calculated from the ratio of
number of symptomatic cases newly diagnosed per 100,000 inhabitants per year during the period 2007–2019 in
Aragon (Spain).
Results: 50 (27.9%) incident cases of HD (CAG repeat length ≥ 36) were identified from a total of 179 persons
studied. The remaining 129/179 (72.1%) were HD negative (CAG repeat length < 36). 29 (58.0%) females and 21(42.0%) males were confirmed as HD cases. The overall incidence was 0.648 per 100,000 patient-years. 11/50
positive HD cases (22.0%) were identified by performing a predictive test, without clinical symptoms. The minimum
number of CAG repeats found was 9 and the most common CAG length among HD negative individuals was 16.
Conclusions: Our incidence lied within the range reported for other Caucasian populations. Implementation of
new techniques has allowed to determine the exact number of CAG repeats, which is especially important inpatients with triplet expansions in an HD intermediate and/or incomplete penetrance allele, both in diagnostic,
predictive and prenatal tests.
Keywords: Huntington disease, HTT gene, Incomplete penetrance alleles, Intermediate alleles

Background
Huntington’s disease (HD) is a progressive neurodegenerative disorder caused by the anomalous expansion ofpolyglutamine repeats (encoded by the trinucleotide
CAG, Cytosine-Adenine-Guanine) in the first exon of
the huntingtin gene (HTT, also known as HD or IT15gene), located in the short arm of chromosome 4(4p16.3) (MIM#143100) [1].
* Correspondence: raquellahoz17@gmail.com
Departamento de Genética. Servicio de Bioquímica Clínica, Hospital
Universitario Miguel Servet. C/ Padre Arrupe, s/n. Consultas Externas. Planta
3ª. 50009, Zaragoza, Spain

HD is an autosomal dominant disease, so the risk thateach child of an affected individual inherits the mutation
is 50%. On the other hand, earlier age of onset in the offspring will depend on the size of the CAGn fragmentthat it inherits, which tends to increase due to thephenomenon of clinical anticipation [2].
There is an inverse relationship between the number
of CAG repeats and the age of onset of symptoms [3, 4].
In addition, the number of CAG repeats is also related
to the age of death [5] and the rate of disease progression [6]. The repeated CAG trinucleotide in the general
population is highly polymorphic, with sizes varying© The Author(s). 2020 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License,
which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give
appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate ifchanges were made. The images or other third party material in this article are included in the article's Creative Commonslicence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commonslicence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain
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The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to thedata made available in this article, unless otherwise stated in a credit line to the data.

Sienes Bailo et al. BMC Medical Genetics(2020) 21:233

between 6 and 26 repeats. However, the diagnostic confirmation of HD involves identifying the presence of mutated alleles of the HTT gene with more than 36 repeats[7]. Alleles with 27–35 CAGs are called intermediate alleles (IAs) and are genetically unstable, being able to be
amplified to pathological sizes in following generations
through the paternal line, and exceptionally, through thematernal [8–11]. Finally, alleles with 36–39 repeats are
related to an incomplete penetrance that is often associated with a later onset age and a slower disease progression rate. Those with 40 or more repeats show complete
penetrance and inevitably lead to the clinical manifestation of HD [12, 13].
Some cases of HD occur sporadically in the population, making very difficult the precise estimate of the
new mutation rate for a late onset disorder such as HD.
It has been determined that 3% of affected patients may
be the result of an expansion of the CAG triplet of an
IA [14]. Thus, it is believed that the existence of thesealleles acts as a reservoir from which de novo mutations
arise in several generations [15].
In this way, the genetic study can be requested for
confirmation of the clinical diagnosis of suspicion oras a presymptomatic or predictive diagnosis. There isalso the possibility of prenatal diagnosis when one ofthe parents is a carrier of a pathological or unstable
expansion, or even select healthy embryos in vitro
for implantation (preimplantation genetic diagnosis)[2, 16].
The main objective of this study was to calculate
the incidence of HD in Aragon population (Spain),
who have been referred to the Clinical Genetics Section of the Hospital Universitario Miguel Servet(HUMS), reference centre in Aragon for the genetic
study of HD, during the period 2007–2019, according
to sex, clinical symptomatology and age at referral.
Other objectives were to estimate the new mutation
rate, to classify the patients diagnosed according to
the CAG triplet repeats and to identify the allele in
the polymorphic normal range that was most common. The number of cases per family were analysed
to assess the paternal or maternal transmission ofHD.

Methods
Subjects/patients

It was a descriptive retrospective study in which thestudy group comprised a total of 179 individuals from
Aragon, who have been referred during the period
January 2007–December 2019 to the Clinical Genetics
Section of the HUMS, Zaragoza, Spain, for diagnostic or
predictive testing for HD. Aragon is an autonomous
community located in north-eastern Spain with a population of around 1.3 million with over half of it living inPage 2 of 8

its capital city, Zaragoza. The area of Aragon represents
a 9.43% of the surface of Spain, being thus the fourth autonomous community in size.
To carry out this study requires the presence ofneurological and/or psychiatric symptoms compatible
with the disease, or a family history information on
HD that justifies it. A family history was considered
positive when at least one other family member has
clinical or genetic diagnosis of HD. A family history
was considered as negative when no other siblings
had signs or symptoms of HD, and the parents wereboth alive and healthy or did not have a neurological/
psychiatric disorder. Demographical data including
age at referral, sex, place of residence and family history are also recorded.
Samples type

The type of primary sample used was peripheral blood
with EDTA from which DNA extraction was performed,
suitable sample for diagnostic studies and presymptomatic/predictive test. In prenatal studies, primary samplesof chorionic villi and amniotic fluid were used.
Methods/techniques

During the period 2007–2012 the determination of the
CAG repeats in these samples was performed using a
conventional polymerase chain reaction (PCR) followedby agarose gel electrophoresis being its main limitation
the inaccuracy of not being able to quantify exactly the
number of repeats of the triplet. From 2013 to 2019,
Adellgene Huntington Disease kit from Blackhills Diagnostic Resources was used in the study of the dynamic
mutation (CAG)n HTT gene (LRG_763t1) (NM_002111.8), located in the locus 4p16.3 by amplifying theregion where is the sequence CAG trinucleotide repeat
and with fluorescence detection. This technology isbased on the amplification of genomic DNA by PCR,
followed by a fluorescent analysis of the size of the PCR
fragments obtained by a genetic analyser, using ABI
3130xl sequencer and GeneMapper 4.0 software whichallows the conversion of said size into the exactly number of CAG repeats using a weight marker, Liz500TM.
The accuracy of sizing is +/− 1 repeat in the normal allele size range and +/− 3 repeats in the expanded allele
size range (> 39 CAG repeats). The laboratory participates successfully in the EMQN (The European Molecular Genetics Quality Network) external quality control
program since 2015, which guarantees the veracity of
the results issued with this technique. Raw data is available as supplementary material.
Study patients were classified according to the number
of CAG repeats they have in the following categories:
normal: 6–26 CAGs; intermediate zone alleles: 27–35
CAGs; incomplete penetrance alleles: 36–39 CAGs;

Sienes Bailo et al. BMC Medical Genetics(2020) 21:233

complete penetrance alleles (adult presentation): 40–59
CAGs; complete penetrance alleles (youth presentation):> 60 CAGs [16]. More detailed clinical and familiar information was obtained of the electronic clinical historyof the patients and their family members to know thetransmission of the disease (paternal or maternal).
The incidence of HD was calculated from the numberof new HD cases diagnosed per 100,000 inhabitants per
year during the period 2007–2019. Binomial confidence
intervals (CI) were also calculated. Annual population
estimates were obtained through the Information Systems Management Department of HUMS belonging to
the Aragonese Health Service.

Page 3 of 8

Table 1 Characteristics of cases undergoing genetic test for HD
CAG repeats ≥36 CAG repeats < 36 Total
Sex
Males

21 (42.0)

58 (45.0)

79 (44.1)

Females29 (58.0)

71 (55.0)

100 (55.9)

Age of testing (years)< 20

1 (2.0)4 (3.1)5 (2.8)20–39

14 (28.0)

24 (18.6)

38 (21.2)

40–49

6 (12.0)

20 (15.5)

26 (14.5)

50–59

9 (18.0)25 (19.4)

34 (19.0)> 60

20 (40.0)56 (43.4)

76 (42.5)

Clinical manifestations

Statistical analyses

Neurological21 (42.0)

69 (53.5)

90 (50.3)

The frequency distribution of the percentages of eachcategory for each qualitative variable was calculated. The
quantitative variables of the study were explored with
the Shapiro-Wilk test (goodness of fit adjustment to anormal distribution) and indicators of central tendency(mean or median) and dispersion (standard deviation or
percentiles) were calculated.
Differences between groups according to sex, age atreferral, genetic testing results and family history wereanalysed by hypothesis contrast test, with comparison of
proportions (χ2) and mean comparisons (Student’s t,Mann-Whitney U, Kruskal-Wallis test) as appropriate.
Data were analysed using the Jamovi statistical softwareand statistical significance was determined at p ≤ 0.05
(two-tailed).

Psychiatric

4 (8.0)2 (1.5)6 (3.3)Mixed

12 (24.0)

20 (15.5)

32 (17.9)

Asymptomatic

13 (26.0)

38 (29.5)

51 (28.5)

37 (74.0)

91 (70.6)

128 (71.5)

Results
A total of 50 (27.9%) incident cases of HD (CAG repeat
length ≥ 36) were identified by genetic testing in theClinical Genetics Section of the HUMS between January
2007 and December 2019 from a total of 179 persons
studied. The remaining 129 of the 179 (72.1%) were HD
negative (CAG repeat length < 36). Samples included inthis database were sent from 19 different primary healthcare centers and hospitals in the Autonomous Community of Aragon. Demographic and clinical characteristics
are shown in Table 1.
According to the data, 29 (58.0%) females and 21(42.0%) males were confirmed as HD cases. Of these,
42 (84.0%) reported a suspected or genetically confirmed positive family history, in three cases (6.0%)
family history was negative and in the remaining five(10.0%) it was impossible to determine the family or
sporadic origin of HD. New mutation events were
genetically proven in one case, estimating that theminimum new mutation rate for HD in our population is 2.0%.
Although the change in the diagnostic technique that
occurred between the periods 2007–2012 and 2013–

Types of testing
Diagnostic
Predictive

11 (22.0)

35 (27.1)

46 (25.7)

Prenatal

2 (4.0)3 (2.3)5 (2.8)Peripheral blood 48 (96.0)

126 (97.7)

174 (97.2)

Amniotic fluid

1 (2.0)1 (0.8)2 (1.1)

Chorionic villi

1 (2.0)2 (1.5)3 (1.7)

Homozygous

0 (0.0)47 (36.4)

47 (26.3)

Heterozygous

50 (100.0)

82 (63.6)

132 (73.7)

SampleZygosity

Demographic, clinical characteristics and genetic test results expressed interms of absolute and percentage relative frequencies: N (%)

2019 has allowed an increase in the number of predictive tests performed (15 (19.5%) v 33 (31.7%) predictivetest), this fact has not modified significantly the numberof positive cases/year (χ2 = 19.2; p = 0.116). However, it
is important to note that 11 (22.0%) of the 50 positive
HD cases were identified by performing a predictive
test, in the absence of any evident clinical symptom.
Number of positive and negative genetic test results for
HD per year between 2007 and 2019 is illustrated inFig. 1. Total population served by the Clinical Genetics
Section of the HUMS during the full period was 593,
387 person-years on average, resulting in an incidence
rate of HD of 0.648 (95% CI 0.587 to 0.709) per 100,000 patient-years, calculated based on the number of
newly diagnosed cases by genetic testing. Positive
cases/year for females (2.23; 95% CI 1.63 to 2.83) andmales (1.62; 95% CI 0.86 to 2.38) were similar, with nostatistically significant differences between them (χ2 =0.0761; p = 0.783).

Sienes Bailo et al. BMC Medical Genetics(2020) 21:233

Page 4 of 8

Fig. 1 Number of positive and negative genetic test results for HD per year (2007–2019)

The mean age of testing was 52.2 years (SD 18.3; range
19–91 years) and more than 43.0% of all HD cases werediagnosed with more than 59 years. Besides, it was similar in females (mean 50.3; SD 19.0; range 19–90 years)
and males (mean 54.7; SD 17.3; range 22–91 years) (U =3894; p = 0.700). In addition, age at referral do not show
any significant difference over the period of the study(K = 20.8; p = 0.077). Among cases with negative family
history, the mean age at referral was 56.1 (SD 8.1) years,
not significantly different from 48.4 (SD 17.0) years on
average obtained in cases with positive family history(U = 47.5; p = 0.495).
The number of CAG repeats in the expanded allele
ranged from 37 to 65 with a mean size of 41.8 (SD 5.8)
repeats. In 32 (14 paternal and 18 maternal) of the 42
cases with positive family history, the type of inheritancewas determined. No significant differences were foundbetween the number of CAG repeats in the expanded allele and the paternal or maternal inheritance (U = 114;
p = 0.648). The minimum number of CAG repeats foundin the study population was 9 and the most common
CAG length among HD negative individuals was 16.

D were prenatal tests performed on a 33 asymptomaticand 30-year-old symptomatic pregnant women, carriersof an incomplete penetrance allele. Fetus C was HD
negative while D was in the incomplete penetrance
range and the pregnancy was interrupted. Finally, case E
was a prenatal test performed on a 33-year-old IVF
pregnant woman diagnosed with HD at the age of 30
which resulted in two alleles in normal range.

Clinical and family history details of patients in whom aprenatal test was performed

In Patient 2, as described in Table 3, a true new mutation event was highly suspected by the identification of
an intermediate allele in three asymptomatic first-degree
relatives. In this case, expansion of an intermediate allele, paternally inherited, was demonstrated confirming a
new mutation event.
There were two symptomatic women without family
history of HD that could be considered possible de novo
mutations, as additional DNA molecular analysis could
not be carried out in their relatives.

The genetic test results for HD, sample type and family
history details of the five prenatal tests performed aresummarized in Table 2.
Case A and B are both descendants of a 33-year-old
asymptomatic female who was a carrier of an incomplete
penetrance allele. Fetus A had an incomplete penetrance
allele while B had a normal and an intermediate allele.
None of these pregnancies were interrupted. Case C andClinical and family history of patients with CAG repeats in
the intermediate range

Genetic test results for HD and family history details of
the seven tests performed are summarized in Table 3.
Frequency of IA in our population (7/308 normal chromosomes) was 2.3%. Despite not being described the relationship between the presence of symptoms inindividuals with alleles in the intermediate range, 42.3%
of the patients with IAs were genetically studied due totheir clinical context with different symptoms of HD.
Clinical and family history details of patients with denovo mutations

Sienes Bailo et al. BMC Medical Genetics(2020) 21:233

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Table 2 Prenatal test cases
Cases

CAG repeats

CAG repeats of parent at risk for transmitting

A1

A2

A1

A2

Sampletype

Family history

A

14

38

13

38

CV

HD confirmed mother, aunt and grandfather

B

13

32

C

15

22

CV

Brother of A

22

39

AF

HD confirmed mother and grandfather

D

16

E

15

37

18

37

AF

HD confirmed mother and grandfather

22

22

47

CV

HD confirmed mother, grandfather and great aunt

Distribution of cases by genetic results, sample type and family history in prenatal test performed
A1 allele, A2 allele 2, CV chorionic villi, AF amniotic fluid

Discussion
More than 43.0% of all HD cases were diagnosed withmore than 59 years, being the mean age of testing 52.2
years-old, which confirms the late onset of the HD [2,16], with no differences between men and women.
Based on the total population served by the Clinical
Genetics Section of the HUMS the overall incidence was
0.648 per 100,000 patient-years, no differences were
found between men and women. In addition, taking into
account the different study periods, similar values wereobtained (0.668 in 2007–2012 and 0.631 in 2013–2019).
A few years earlier, in the Basque country and Navarre
in Spain (geographical areas adjacent to our community), Ramos-Arroyo et al. determined an incidence of
0.48 and 0.46 per 100,000 patient-years respectively [14],being lower than the incidence found in our population.
In other European countries, the incidence rates calculated were the following, all of them per 100,000 personyears: 0.3 in the province of Ferrara, northern Italy(1990–2009) [17]; 1.8 in Germany (2015–2016) [18];0.14 in Iceland (1988–2007) [19]; the minimum incidence rate was 0.22, being able to reach 0.44 in Greece(1995–2008) [20]; 0.25 in Cyprus (1984–2015) [21]; and0.72 in United Kingdom (1990–2010) [22]. So, the incidence of HD varies from 0.14 to 1.8 per 100,000 personyears in Europe.
In the population of the United States from 2003 to2016 the incidence rate was 1.22 per 100,000 personyears [23] while in Asian countries it is much lower(0.06 in Korea during 2009–2013 [24] and 0.1 in Taiwan
during 2000–2007 [25]). In their review, Pringsheim
et al. found a mean incidence of 0.4 per 100,000 personyears. Studies including non-Asian population has ahigher incidence (0.1–0.8) than studies with Asian descent populations (0.05–0.1) [26]. No incidence studies
have been found in the African population [27], although the prevalence of HD in sub-Saharan areas andsouthern Africa has been found to be lower than in populations of Caucasian origin [27–29].
The incidence in Aragon is higher than the reported
for other Caucasian populations, although exists an apparent heterogeneity that varies both time and place. Indeed, this heterogeneity appeared in the estimations of
the HD prevalence too, being lower in studies carriedout in Asia and Africa than in those that include predominantly populations of European descent [22, 27,

Table 3 Cases with CAG repeats in the intermediate range (27–35) and their relatives with other CAG repeats
Patient

Sex

CAG repeats

1

Male

17

29

HD confirmed two siblings–

Relatives

Male

19

44

Brother

Depression

Female1744

SisterEssential tremor, bradykinesia, stiffness in the upper right limb

Female1719

Sister–

A1

Family history

Clinical manifestations

A2

2a

Male

16

43

Negative

Generalized dystonia, multiple motor tics

Relatives

Male

19

33

Father–Female1733

Sister

–

Male

17

33

Brother–3

Female18

27

Negative

Generalized choreic movements, muscle contractions

4

Female17

31

HD suspected father and paternal aunts

Depression, anxiety, phobia, chronic stress

5

Male

18

34

Negative

Progressive instability, choreic movements and restlessness

Distribution of cases by genetic results of intermediate allele and family history in predictive and diagnostic test performed. A1 allele, A2 allele 2. aIn this case, theproband (patient 2) presented an allele in the full penetrance range, being his relatives the carriers of the intermediate allele

Sienes Bailo et al. BMC Medical Genetics(2020) 21:233

29]. Differences in diagnostic criteria and case ascertainment could explain this variety. Also, if pre-symptomaticcases were included, to the proportion of individuals
opting for a predictive testing, which would increase thenumber of cases. Since the early 1990s for the HD mutation exists a molecular genetic test for its diagnostic.
This way, depending on its availability in the different
geographical regions, the estimate of new cases mayvary. Finally, some authors have suggested the presenceof modifiers of the expression of the disease (genetic or
environmental) that could play an important role in the
expansions of the HD allele around the world, what
would make the population more or less prone to thedisease [30, 31].
Given that the genetic mutation presumably originatedin north-western Europe [26], it suggests that there
could be a founder effect in Aragon. This would be possible since the incidence is higher in this region compared to other European populations, but more similar
to the Basque Country and Navarra, neighbouring communities also located in north-eastern Spain. However, it
would be necessary to carry out more extensive studies
for its confirmation.
This complex landscape contributes to an uncertain
epidemiology for HD and incidence rates are probably
underestimated despite the progress in the knowledge ofthis disease in recent years [17].
The most common CAG length in our population
among the normal alleles was 16, slightly lower than thelength found in European population with 17 CAGs repeats [7]. In Danish population, this size is the most frequent too [32], as well as for Canadians [33].
The mean size of 17.2 (SD 2.8) repeats for Aragonese
people was similar to that found in European population, being larger compared to the mean size in Asian
countries such as China, Japan and Thailand (16.4, 16.6
and 16.5 repeats respectively) [24]. In Latin America themean varies from 15 to 17.8 repeats, similar to those reported among Caucasians an Asians [34], being 16 themost common repeat among HD negative individuals in
Cuba and 17 in Venezuela [35].
These variations in the normal range of CAG repeats
according to the population studied (greater for countries whose offspring are of Caucasian origin than inthose of Asian origin) correspond to those observed in
the incidence and prevalence of the HD, since both are
directly related [34]. In populations with high prevalenceof HD is higher the number of CAG repeats in normal
chromosomes [35].
Planning a family and prevention of passing on an expanded CAG-repeat to future offspring are the main reasons to apply for prenatal or preimplantation genetic
diagnosis [36]. In our population, four prenatal and one
preimplantation tests were carried out for 4 mothers

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affected of HD. In these cases, the importance of genetic
counselling was revealed, being necessary the translationof the advances in genetic and in-utero techniques into
genetic counselling, taking into account the ethical andscientific principles allowing families to make decisions,
including the interruption of pregnancy [37], as one of
our cases. It is important because younger generations
are likely to be more seriously affected by inheriting a
CAG repeat amplification, so the detection of the disease
and prenatal tests would make it possible to inform andcounsel other family members [14].
The frequency of IA in our population was 2.3%, similar to other general populations being observed in 1–7%,
which can be considered commonly present [36]. The
42.3% of cases in this range presented clinical symptoms.
Some years ago, IAs were not considered associated withsymptomatic HD. Nevertheless, nowadays there is evidence that some individuals with IAs express the diseasephenotype, developing HD-like clinical and neuropathological manifestations, as in our study, being more notable in older patients or in those with greater repeats
within the range 27–35 [8].
When IA has more than 30 repeats, the probability
that an expanded allele could be transmitted to the offspring is higher, being able to reach full penetrance [7].
In addition to the size of CAG, the likelihood of expansion to the disease associated range is highly influenced
by the sex of the transmitting parent, being the majorityof new mutations of paternal transmission [38], althoughalso could be inherit by maternal line [9]. This explains
our only confirmed case of a new mutation in which thefather with 33 CAG repeats has been responsible for thetransmission of the disease to his son (Patient 2) in therange of complete penetration.
This way during meiosis, IA instability results in denovo expansions, an uncommon but well-known sourceof new HD cases in individuals with no family history.
The minimum new mutation rate for HD in our population was 2.0%, below the new mutation rate for theBasque country and Navarre with a minimum of 4.7%
and a potential rate of 8.1% [14], although the new mutation rate for HD is estimated to be 10.0% [39]. In thesepatients, the negative family history of HD observed
could be explained by new mutations, but also it could
be due to misdiagnosis in family members, nonpenetrance or non-paternity [20]. So further studies will
be needed to confirm these findings, including genetic
testing of family members even those asymptomaticones. This fact is important information to take into account when genetic counselling a family with a true denovo HD case [40].
The main limitations of the study were found in thecalculation of the de novo rate of new mutations, sincein two patients their relatives could not be geneticallySienes Bailo et al. BMC Medical Genetics(2020) 21:233

tested. In addition, the association of clinical symptoms
in those patients with IA is controversial. Other diseases
with the same type of symptoms should be studied inorder to be able to firmly affirm that this symptomatology is due to the presence of these alleles. Finally, the incidence has been calculated with reference to the
population that belongs to our health sector, but it may
be neglecting patients who, even belonging to this area,
have carried out the study elsewhere.

Conclusions
Despite being a rare disease, the incidence of HD in our
population can be considered relevant. The implementation of new techniques in the Genetics Laboratory suchas fluorescent PCR has allowed us to determine theexact number of CAG repeats, which is especially important in those patients with triplet expansions in an
HD intermediate and/or incomplete penetrance allele,both in diagnostic, predictive and prenatal tests. In
addition, this technology enables us to have truthful andreliable results in 12–24 h offering an adequate andtimely genetic counselling to the patient and theirrelatives.
Supplementary Information
The online version contains supplementary material available at https://doi.
org/10.1186/s12881-020-01174-z.
Additional file 1: Suplementary table 1. Datasets generated andanalysed.
Abbreviations
CAG: Cytosine-Adenine-Guanine; CEICA: Comité de Ética de la Investigaciónde la Comunidad de Aragón; CI: Confidence interval; EMQN: The European
Molecular Genetics Quality Network; HD: Huntington disease; HUMS: Hospital
Universitario Miguel Servet; IAs: Intermediate alleles; PCR: Polymerase chain
reaction; SD: Standard deviation
Acknowledgements
Not applicable.
Authors’ contributions
SIA acquired and analysed the patient data regarding the genetic analysis ofHuntington disease. PSB and RL analysed the database and interpreted it.
Also, they were the major contributors in writing the manuscript. JPSM and
SIA substantively revised the work. All authors read and approved the final
manuscript.
Funding
This research did not receive any specific grant from funding agencies in the
public, commercial, or not-for-profit sectors.
Availability of data and materials
The datasets generated and analysed during the current study are available
as Supplementary Material.
The website https://www.ncbi.nlm.nih.gov/nuccore/NM_002111 was
consulted to obtain reference information about the HTT gene.
Ethics approval and consent to participate
This research project with code PI19/403 was approved by the Research
Ethics Committee of the Community of Aragon (CEICA, Comité de Ética dela Investigación de la Comunidad de Aragón). Administrative permissions

Page 7 of 8

were acquired to access the clinical/personal patient data used and were
granted by the head of the Clinical Documentation and Archives Unit, thePresident of the Research and Innovation Commission and the Director of
the hospital. This is a retrospective study and the data were collected underclinical practice conditions, with no additional procedures being performedon patients beyond those necessary for the study of the pathology, so it was
not necessary for the patients to sign an additional consent as authorized by
the Ethics Committee. Previously, the patients had already signed a written
informed consent for the genetic study and in the case of any participant
under the age of 16 it was obtained from the parents or legal guardians.
Consent for publication
Not applicable.
Competing interests
The authors declare that they have no competing interests.
Received: 22 May 2020 Accepted: 12 November 2020

References
1. Pandey M, Rajamma U. Huntington’s disease: the coming of age. J Genet.
2018;97:649–64.
2. Ramos Arroyo MA, Trujillo-Tiebas MJ, Milá M. Recomendaciones de buena
práctica para el diagnóstico genético de la enfermedad de Huntington.
Med Clínica. 2012;138:584–8.
3. Duyao M, Ambrose C, Myers R, Novelletto A, Persichetti F, Frontali M, et al.
Trinucleotide repeat length instability and age of onset in Huntington’sdisease. Nat Genet. 1993;4:387–92.
4. Andresen JM, Gayán J, Djoussé L, Roberts S, Brocklebank D, Cherny SS, et al.
The relationship between CAG repeat length and age of onset differs for
Huntington’s disease patients with juvenile onset or adult onset. Ann Hum
Genet. 2007;71:295–301.
5. Persichetti F, Srinidhi J, Kanaley L, Ge P, Myers RH, D’Arrigo K, et al.
Huntington’s disease CAG trinucleotide repeats in pathologically confirmed
post-mortem brains. Neurobiol Dis. 1994;1:159–66.
6. Rosenblatt A, Liang K-Y, Zhou H, Abbott MH, Gourley LM, Margolis RL, et al.
The association of CAG repeat length with clinical progression in
Huntington disease. Neurology. 2006;66:1016–20.
7. Semaka A, Kay C, Doty C, Collins JA, Bijlsma EK, Richards F, et al. CAG sizespecific risk estimates for intermediate allele repeat instability in Huntington
disease. J Med Genet. 2013;50:696–703.
8. Cubo E, Ramos-Arroyo MA, Martinez-Horta S, Martínez-Descalls A, Calvo S,
Gil-Polo C, et al. Clinical manifestations of intermediate allele carriers in
Huntington disease. Neurology. 2016;87:571–8.
9. Semaka A, Kay C, Belfroid RDM, Bijlsma EK, Losekoot M, van Langen IM,
et al. A new mutation for Huntington disease following maternaltransmission of an intermediate allele. Eur J Med Genet. 2015;58:28–30.
10. Groen JL, de Bie RMA, Foncke EMJ, Roos RAC, Leenders KL, Tijssen MAJ.
Late-onset Huntington disease with intermediate CAG repeats: true or false?
J Neurol Neurosurg Psychiatry. 2010;81:228–30.
11. Kremer B, Almqvist E, Theilmann J, Spence N, Telenius H, Goldberg YP, et al.
Sex-dependent mechanisms for expansions and contractions of the CAG
repeat on affected Huntington disease chromosomes. Am J Hum Genet.
1995;57:343–50.
12. McNeil SM, Novelletto A, Srinidhi J, Barnes G, Kornbluth I, Altherr MR, et al.
Reduced penetrance of the Huntington’s disease mutation. Hum Mol
Genet. 1997;6:775–9.
13. Potter NT, Spector EB, Prior TW. Technical standards and guidelines for
Huntington disease testing. Genet Med Off J Am Coll Med Genet. 2004;6:
61–5.
14. Ramos-Arroyo M, Moreno S, Valiente A. Incidence and mutation rates ofHuntington’s disease in Spain: experience of 9 years of direct genetic
testing. J Neurol Neurosurg Psychiatry. 2005;76:337–42.
15. García-Planells J, Burguera JA, Solís P, Millán JM, Ginestar D, Palau F, et al.
Ancient origin of the CAG expansion causing Huntington disease in a
Spanish population. Hum Mutat. 2005;25:453–9.
16. Losekoot M, van Belzen MJ, Seneca S, Bauer P, Stenhouse SAR, Barton DE.
EMQN/CMGS best practice guidelines for the molecular genetic testing of
Huntington disease. Eur J Hum Genet. 2013;21:480–6.

Sienes Bailo et al. BMC Medical Genetics(2020) 21:233

17. Carrassi E, Pugliatti M, Govoni V, Sensi M, Casetta I, Granieri E.
Epidemiological study of Huntington’s disease in the province of Ferrara,
Italy. Neuroepidemiology. 2017;49:18–23.
18. Ohlmeier C, Saum K-U, Galetzka W, Beier D, Gothe H. Epidemiology andhealth care utilization of patients suffering from Huntington’s disease inGermany: real world evidence based on German claims data. BMC Neurol.
2019;19. https://doi.org/10.1186/s12883-019-1556-3.
19. Sveinsson O, Halldórsson S, Olafsson E. An unusually low prevalence ofHuntington’s disease in Iceland. Eur Neurol. 2012;68:48–51.
20. Panas M, Karadima G, Vassos E, Kalfakis N, Kladi A, Christodoulou K, et al.
Huntington’s disease in Greece: the experience of 14 years. Clin Genet.
2011;80:586–90.
21. Demetriou CA, Heraclides A, Salafori C, Tanteles GA, Christodoulou K,
Christou Y, et al. Epidemiology of Huntington disease in Cyprus: a 20-year
retrospective study. Clin Genet. 2018;93:656–64.
22. Wexler NS, Collett L, Wexler AR, Rawlins MD, Tabrizi SJ, Douglas I, et al.
Incidence of adult Huntington’s disease in the UK: a UK-based primary carestudy and a systematic review. BMJ Open. 2016;6:e009070.
23. Bruzelius E, Scarpa J, Zhao Y, Basu S, Faghmous JH, Baum A. Huntington’sdisease in the United States: variation by demographic and socioeconomic
factors. Mov Disord. 2019;34:858–65.
24. Kim HS, Lyoo CH, Lee PH, Kim SJ, Park MY, Ma H-I, et al. Current status of
Huntington’s disease in Korea: a Nationwide survey and National Registry
Analysis. J Mov Disord. 2015;8:14–20.
25. Chen Y-Y, Lai C-H. Nationwide population-based epidemiologic study of
Huntington’s disease in Taiwan. Neuroepidemiology. 2010;35:250–4.
26. Pringsheim T, Wiltshire K, Day L, Dykeman J, Steeves T, Jette N. The
incidence and prevalence of Huntington’s disease: a systematic review andmeta-analysis. Mov Disord. 2012;27:1083–91.
27. Lekoubou A, Echouffo-Tcheugui JB, Kengne AP. Epidemiology ofneurodegenerative diseases in sub-Saharan Africa: a systematic review. BMC
Public Health. 2014;14:653.
28. Baine F, Kay C, Ketelaar M, Collins J, Semaka A, Doty C, et al. Huntington
disease in the south African population occurs on diverse and ethnicallydistinct genetic haplotypes. Eur J Hum Genet. 2013;21:1120–7.
29. Baine F, Krause A, Greenberg L. The frequency of Huntington disease andHuntington disease-like 2 in the south African population.
Neuroepidemiology. 2016;46:198–202.
30. Xu M, Wu Z-Y. Huntington Disease in Asia. Chin Med J (Engl). 2015;128:
1815–9.
31. Warby SC, Visscher H, Collins JA, Doty CN, Carter C, Butland SL, et al. HTT
haplotypes contribute to differences in Huntington disease prevalence
between Europe and East Asia. Eur J Hum Genet. 2011;19:561–6.
32. Gilling M, Budtz-Jørgensen E, Boonen SE, Lildballe D, Bojesen A, Hertz JM,
et al. The Danish HD registry-a nationwide family registry of HD families inDenmark. Clin Genet. 2017;92:338–41.
33. Semaka A, Kay C, Doty CN, Collins JA, Tam N, Hayden MR. High frequency
of intermediate alleles on Huntington disease-associated haplotypes in
British Columbia’s general population. Am J Med Genet Part B
Neuropsychiatr Genet Off Publ Int Soc Psychiatr Genet. 2013;162B:864–71.
34. Castilhos RM, Augustin MC, Santos JA, Perandones C, Saraiva-Pereira ML,
Jardim LB. Genetic aspects of Huntington’s disease in Latin America. A
systematic review. Clin Genet. 2016;89:295–303.
35. Apolinário TA, Paiva CLA, Agostinho LA. REVIEW-ARTICLE intermediate
alleles of Huntington’s disease HTT gene in different populations
worldwide: a systematic review. Genet Mol Res GMR. 2017;16.
36. Tibben A, Dondorp WJ, de Wert GM, de Die-Smulders CE, Losekoot M,
Bijlsma EK. Risk assessment for Huntington’s disease for (future) offspring
requires offering Preconceptional CAG analysis to both partners. J Huntingt
Dis. 2019;8:71–8.
37. Migliore S, Jankovic J, Squitieri F. Genetic counseling in Huntington’sdisease: potential new challenges on horizon? Front Neurol. 2019;10.
https://doi.org/10.3389/fneur.2019.00453.
38. Semaka A, Hayden MR. Evidence-based genetic counselling implications for
Huntington disease intermediate allele predictive test results. Clin Genet.
2014;85:303–11.
39. Savitt D, Jankovic J. Clinical phenotype in carriers of intermediate alleles in
the huntingtin gene. J Neurol Sci. 2019;402:57–61.
40. Houge G, Bruland O, Bjørnevoll I, Hayden MR, Semaka A. De novo
Huntington disease caused by 26-44 CAG repeat expansion on a low-risk
haplotype. Neurology. 2013;81:1099–100.

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